Acute slice preparation
Sodium pentobarbital (50 mg/kg in a 1 mL/kg volume) was used for deep
anesthesia; animals were then decapitated under its effects. Once
removed from the skulls, brains were placed into a frosty sucrose
solution containing (in mM): 210 sucrose, 2.8 KCl, 2
MgSO4, 1.25 Na2HPO4, 25
NaHCO3, 1 MgCl2, 1
CaCl2, and 10 D-glucose. The sucrose solution was
continuously bubbled with a carbogen mixture (95%
O2/5% CO2). Tissue blocks of cerebral
hemispheres containing the hippocampus and surrounding areas were sliced
at 385 µm thickness in the transversal plane using a vibrating tissue
slicer (Leica VT1000S; Nusschloc, Germany). Next, the fresh slices were
stabilized at 34° C for 25 to 30 min in an artificial cerebrospinal
fluid solution (ACSF; pH ≈ 7.30–7.35) containing (in mM): 125 NaCl, 2.5
KCl, 1.25 Na2HPO4, 25
NaHCO3, 4 MgCl2, 1
CaCl2, and 10 D-glucose. Next, the slices were kept at
room temperature for at least 1 hour before any experimental procedure
was performed. An individual slice was transferred to a submerged
chamber (total volume: 400 µL). The slice was continuously perfused with
modified ACSF at a rate of 2.2–2.5 mL/min, with the help of a
peristaltic pump (Sci Q400, Watson-Marlow, Wilmington, MA, USA). The
modified ACSF (with continuous carbogen bubbling) included the following
components (in mM): 125 NaCl, 2.5 KCl, 1.25
Na2HPO4, 25 NaHCO3, 1.5
MgCl2, 2.5 CaCl2, and 10 D-glucose. The
recordings were performed at 32.5 ±1 °C.